IRREVERSIBLE INACTIVATION OF PORCINE PEPSIN BY ϒ-IRRADIATION IN AQUEOUS SOLUTION

Open Access
- Author:
- Trinh, Eric Quang Dao
- Area of Honors:
- Chemistry
- Degree:
- Bachelor of Science
- Document Type:
- Thesis
- Thesis Supervisors:
- Bratoljub Milosavljevic, Thesis Supervisor
Bratoljub Milosavljevic, Thesis Supervisor
Karl Todd Mueller, Thesis Honors Advisor
John B Asbury, Faculty Reader - Keywords:
- pepsin
radiation
activity
ovalbumin
sds-page
agglomeration
scission - Abstract:
- The effects of ϒ-irradiation on the activity and structure of pepsin from porcine gastric mucosa were studied. 1% w/v aqueous enzyme solutions were irradiated with doses of 1, 2.5, 5, and 10 kGy; pristine enzyme was used as a control for normal enzyme behavior. Purified, lyophilized ovalbumin from Gallus gallus eggs was used as the substrate for proteolytic cleavage. The activity of pepsin was measured by the appearance of soluble cleavage products determined by a spectrophotometric assay for primary amines; measurements were compared to standard curves created using pure leucine and valine solutions. Nominal changes in the structure of the enzyme were analyzed using sodium dodecyl sulfate gel electrophoresis. Measurement of product formation during the time-resolved digestions of ovalbumin performed with the irradiated enzyme solutions indicated a positive correlation between loss of enzyme activity and increasingly greater doses of radiation. The radical reactions and scission induced by ϒ-radiation, which have been studied in previous experiments, appear to have the ability to irreversibly inactivate pepsin. These results underscore the dependence of protein function on structure, and immediately suggest several different ways through which radical reactions may inactivate an enzyme's catalytic ability. Further studies following up on these results could yield a better understanding of the exact nature of how radiation affects more complicated in vivo systems.